Drug resistance, including resistance to topoisomerase II (topo II) inhibitors, is a major obstacle in the treatment of multiple myeloma. Topo II poisons that are used in the treatment of multiple myeloma include doxorubicin and etoposide (VP-16). Several mechanisms of resistance to topo IIα inhibitors have been described [Rasheed Z A, et al., Oncogene 2003; 22:7296-304; Oloumi A, et al., Cancer Res 2000; 60:5747-53]. Cell adhesion-mediated drug resistance and stromal cell adherence are important parameters in the local bone marrow environment in patients with multiple myeloma and appear to be major determinants of drug resistance [Hazlehurst L A, et al., Blood 2001; 98:1897-903; Hazlehurst L A, et al., Cancer Res 2006; 66:2338-45].
Human multiple myeloma cell density is a determinant of sensitivity to topo II inhibitors [Valkov N I, et al., Br J Haematol 2000; 108:331-45; Turner J G, et al., J Cell Sci 2004; 117:3061-71]. At increased cell densities, a considerable fraction of nuclear topo IIα is exported to the cytoplasm (>90%), resulting in reduced sensitivity to VP-16 and doxorubicin. This appears to occur both in human myeloma cell lines and in CD138+ cells isolated from patients with multiple myeloma. Nuclear export of topo IIα may contribute to drug resistance, and that resistance may not be due to differences in drug uptake, cell cycle, or cellular topo IIα protein levels [Valkov N I, et al., Br J Haematol 2000; 108:331-45; Turner J G, et al., J Cell Sci 2004; 117:3061-71]. The nuclear export signals for topo IIα are found at amino acids 1,017 to 1,028 and 1,054 to 1,066 [Turner J G, et al., J Cell Sci 2004; 117:3061-71]. Export can be blocked by treatment of the cells with leptomycin B, indicating that a CRM1-dependent pathway mediates export [Turner J G, et al., J Cell Sci 2004; 117:3061-71]. In the present study, we show that inhibition of CRM1-mediated export of topo IIα may render myeloma cells both in vitro and ex vivo more sensitive to topo II-targeted chemotherapy.
Use of CRM1 inhibition in cancer therapy has met with limited success. The first CRM1 inhibitor, leptomycin B, was found to efficiently inhibit nuclear export. However, leptomycin was found to have acute relative toxicities both in a human phase I trial [Newlands E S, et al., Br J Cancer 1996; 74:648-9] and in vitro. Leptomycin B in vitro studies found acute toxicity at concentrations<5 nmol/L for 1 hour. Therefore, in this study, we used the CRM1 inhibitor ratjadone C [Turner, J et al., Cancer Research (2009)69(17):6899-905; Falini B, et al., Blood (2006)107:4514-23]. Ratjadone C has been found to inhibit nuclear export without producing apoptosis or necrosis at concentrations up to 300 nmol/L for 48 h in an in vitro assay. However, ratjadone C prevents nuclear export of topo IIα; in this study, we show that ratjadone C also acutely sensitizes myeloma cells to the topo II inhibitors doxorubicin and VP-16. Additional low-toxicity CRM1 inhibitors are also being investigated by other laboratories and may become available for preclinical studies.